Introduction: Resistance to extended-spectrum cephalosporins in E. coli can be meditated by mobile AmpC-type b-lactamases and over-expression of native AmpC enzymes normally repressed. Herein, we describe a full genetic characterisation of AmpC-mediated resistance and report novel insertion promoter sequences.

Methods: The Norwegian Reference Centre for Detection of Antimicrobial Resistance (K-res) received 414 clinical isolates of E. coli from different Norwegian laboratories with a reduced susceptibility to third-generation cephalosporins, over a 2-year period 20032005. Of these, 23 isolates showed an AmpC-mediated phenotype using substrate profiling. Strains were molecularly characterised and typed by IEF and PFGE. AmpC-type b-lactamase genes and adjacent sequences were analysed using PCR, sequence analysis, endonuclease plasmid analysis and hybridisation.

Results: The 23 isolates all possessed pI values of pI 9.09.2 indicative of AmpC expression. Multiplex blaampC-PCR and southern blot analysis of S1 nuclease digested total DNA separated by PFGE, suggests that 10 of the 23 E. coli isolates have the blaampC gene located on plasmids. Sequencing analysis showed that 9 of these had the blaCMY-2 gene, while one strain had the blaCMY-7 gene. All plasmid-mediated blaampC genes were found to be linked to an insertion sequence ISEcp1-like element by an ISEcp1-blaampC gene linkage PCR. The bla CMY-2 genes possessed blc, sugE and dsbC downstream suggesting that they belong to Type 1 and Type 2. The 13 non-plasmid mediated blaampC strains, revealed promoter and attenuator alterations compared to E. coli wild-type promoters. Two blaampC promoter regions were interrupted by the insertion sequence ISEc10 of the IS21-family. PFGE analysis suggests that the majority of the 23 E. coli isolates are unrelated.

Conclusion: These findings suggest there are two different populations of AmpC-type resistant E. coli clinical isolates in Norway  plasmid-mediated blaCMY genes and cognate AmpC promoter mutations. The genetic context of the blaCMY gene suggests they are identical to international strains.